Survival of Anaplasma marginale in Aedes albopictus cells

Bovine red blood cells infected with Anaplasma marginale were introduced into cell line cultures from the mosquito Aedes albopictus. The cultured mosquito cells phagocytized both infected and uninfected red blood cells. The parasite remained viable, as determined by calf inoculations, for 21 days.     

Does transovarial transmission of dengue virus occur in Malaysian Aedes aegypti and Aedes albopictus?

PURPOSE: We assessed the relative difficulty with which radical perineal and retropubic prostatectomy operations are learned. MATERIALS AND METHODS: The first 10 radical perineal and retropubic prostatectomies performed by 6 graduating urology residents were reviewed for patient and tumor characteristics, duration of surgery and hospitalization, and complication rates. RESULTS: A total of 120 patients was reviewed. Mean hospital stay was 2.5 days less after radical perineal prostatectomy, and estimated blood loss, number of transfusions and complication rates were also less for this procedure. All parameters used to estimate the outcome of the procedure indicated that radical perineal prostatectomy was learned more quickly than radical retropubic prostatectomy. CONCLUSIONS: In a residency training program radical perineal prostatectomy was learned at least as easily as retropubic prostatectomy. Due to expanding indications for this procedure, these findings should encourage urological surgeons to develop their skill with this approach.     

Aedes albopictus (Skuse) (Diptera, Culicidae) in eastern Veneto

A survey was carried out in autumn 1992 and spring-autumn 1993 to study the seasonal distribution and the dispersal of Aedes albopictus (Skuse) in Portogruaro (province of Venice; North-Eastern Italy). Larvae and pupae were found from May through September; the larvae hatching from eggs after the end of September seemed unable to complete development. The adults bit until October. The mosquito spread in new areas of Portogruaro town, even after insecticidal treatments. Private areas seemed to play an important role in the preservation of the infestation, as they were not treated by the public pest control service.     

Susceptibility of three laboratory strains of Aedes albopictus (Diptera: Culicidae) to Japanese encephalitis virus from Taiwan

An 11-year-old boy developed florid reactive periostitis several years after minor trauma. The symptoms responded initially to antibiotics, but after cessation, rapidly recurred and progressed, requiring a ray amputation to relieve the pain and to achieve a functional hand. The reactive periostitis affected the volar aspect of two adjacent phalanges with sparing of the intervening joint, confirming that this is a reactive process rather than a benign neoplasm.     

Breeding habitats of Aedes aegypti (L) and Aedes. albopictus (Skuse) in villages of Barru, South Sulawesi, Indonesia

The breeding habitats of the dengue vector, Aedes aegypti and Aedes albopictus, were studied using larval collection method inside and outside houses in 6 villages of Barru, South Sulawesi, Indonesia from July 1994 to August 1995. Aedes aegypti was the dominant species, being abundant indoors especially in the coastal areas. Aedes albopictus was breeding primarily in outdoor containers in the hill and mountain areas. Earthen jar was the most common breeding habitat of Aedes aegypti in all villages surveyed. Drum can was the most common outdoor breeding habitat of Aedes albopictus in the hill and mountain areas. The high Breteau indices of Aedes aegypti and Aedes albopictus suggests that these species may play an important role in the transmission of dengue hemorrhagic fever in Barru where epidemics of the fever occur occasionally.     

Structural localization of the E3 glycoprotein in attenuated Sindbis virus mutants

We have determined the three-dimensional structures of the wild-type Sindbis virus and two of its mutants that retain the E3 sequence within PE2. Using difference imaging between these mutants and the wild-type virus, we have assigned a location for the 64-amino-acid sequence corresponding to E3 in the mutant spike complex. In the wild-type virus, the spike is composed of an E1-E2 heterotrimer. The E3 protein was found to protrude midway between the center of the spike complex and the tips. Based on these results and the work of others, we propose a distribution for the functional domains of the spike proteins within the structure of wild-type Sindbis virus. Within the structure of the virus, the E1 domains form the central portion of the spike complex, while the tips are formed by the E2 domains that flare out from the center of the complex. The structural similarity between these Sindbis virus mutants and Ross River virus suggests that E3 may also be present in the latter, which is also a member of the Alphavirus genus.     

Ultrastructure and immuno-cytochemistry of BHK-21 cells infected with a modified Bucyrus strain of equine arteritis virus

Morphogenesis of a modified Bucyrus strain of equine arteritis virus (EAV) in BHK-21 cells was studied. Bacillary tubules were first detected in the cytoplasm 8 h after infection, and mature virions 79 to 122 nm in diameter, 101 nm on average, were mostly observed in the cisternae of the rough endoplasmic reticulum (RER) at 12 h or later. They had isometrical cores and morphological subunits in the outer layer. Budding occurred from the RER and the outer nuclear membrane, but not from the cell surface. Structural linkage was detected between the tubule and the virus core. Aberrant strands were occasionally demonstrated within the nucleus 12 h after infection, and immunofluorescence and immunogold labeling revealed viral antigen also in the nucleus.     

Rapid selection in modified BHK-21 cells of a foot-and-mouth disease virus variant showing alterations in cell tropism

With persistent foot-and-mouth disease virus (FMDV) in BHK-21 cells, there is coevolution of the cells and the resident virus; the virulence of the virus for the parental BHK-21 cells is gradually increased, and the cells become partially resistant to FMDV. Here we report that variants of FMDV C3Arg/85 were selected in a single infection of partially resistant BHK-21 cells (termed BHK-Rb cells). Indirect immunofluorescence showed that the BHK-Rb cell population was heterogeneous with regard to susceptibility to C3Arg/85 infection. Infection of BHK-Rb cells with C3Arg/85 resulted in an early phase of partial cytopathology which was followed at 6 to 10 days postinfection by the shedding of mutant FMDVs, termed C3-Rb. The selected C3-Rb variants showed increased virulence for BHK-21 cells, were able to overcome the resistance of modified BHK-21 cells to infection, and had acquired the ability to bind heparin and to infect wild-type Chinese hamster ovary (CHO) cells. A comparison of the genomic sequences of the parental and modified viruses revealed only two amino acid differences, located at the surface of the particle, at the fivefold axis of the viral capsid (Asp-9-->Ala in VP3 and either Gly-110-->Arg or His-108-->Arg in VP1). The same phenotypic and genotypic modifications occurred in a highly reproducible manner; they were seen in a number of independent infections of BHK-Rb cells with viral preparation C3Arg/85 or with clones derived from it. Neither amino acid substitutions in other structural or nonstructural proteins nor nucleotide substitutions in regulatory regions were found. These results prove that infection of partially permissive cells can promote the rapid selection of virus variants that show alterations in cell tropism and are highly virulent for the same cells.     

Nutritional reserves of autogenous and anautogenous selected strains of Aedes albopictus (Diptera: Culicidae)

Autogeny in Aedes albopictus (Skuse) females was increased from 5% in wild-type parental stock to 84% by selection through 20 generations and was decreased to 0% by selection through nine generations. Concomitantly, the mean number of eggs per female that matured autogenously increased from 6.4 to 18.3 in sugar-fed mosquitoes that developed eggs. Although the duration of the larval period of the two strains was not different, autogenous females emerged with greater amounts of metabolizable protein and lipid and total available energy reserves than did anautogenous females, but with lesser amounts of glycogen. Dry weights did not differ significantly. The difference in the energy reserves between strains was slightly more than the total energy invested in 20 autogenous eggs. Autogenous females survived 1.3 d longer than did anautogenous females when starved from emergence.     

Laboratory and field evaluations of oviposition responses of Aedes albopictus and Aedes triseriatus (Diptera: Culicidae) to oak leaf infusions

Organic infusions created by fermenting white oak (Quercus alba L.) leaves in water were evaluated as sources of attractant odorants and contact oviposition stimulants for gravid Aedes albopictus (Skuse) and Aedes triseriatus (Say). Infusions were bioassayed in the laboratory by giving single females a choice of ovipositing in 1 container with infusion and 7 containers with water. Ae. albopictus laid significantly more eggs in containers with infusion, regardless of concentration (dilutions ranging from 10 to 100%) or age (fermentation periods of 7, 28, 60 d), than in containers holding water. The largest proportion of eggs (76.8%) was deposited in response to a 60% concentration of 7-d-old infusion. In contrast, Ae. triseriatus exhibited variable oviposition responses but generally deposited the largest number of eggs in only a few concentrations of older age infusions. In binary "sticky screen" bioassays, there was no difference between the numbers of females attracted to infusion or water, indicating that oviposition responses to infusion were mediated by contact chemostimulants rather than by attraction to odorants. Oviposition responses to infusions by field populations of Ae. albopictus and Ae. triseriatus in Raleigh, NC, were evaluated with pairs of oviposition traps, one containing infusion and the other containing water. Generally, Ae. albopictus laid significantly more eggs in ovitraps containing infusion regardless of its age (7, 28, and 60 d old) or the mass of leaves fermented (126 g = 1x or 504 g = 4x) than in water. In contrast, Ae. triseriatus deposited an equivalent number of eggs in traps containing water or 1x, 80% infusion regardless of its age; however, the oviposition response to ovitraps containing 4x, 7-d-old, 50% infusion was significant. Placement of an automobile tire behind an ovitrap did not increase the number of Ae. albopictus eggs laid in ovitraps containing 4x, 7-d-old, 50% infusion or water relative to ovitraps without a tire. Our research indicates that baiting ovitraps with oak leaf infusion would increase the sensitivity of surveillance efforts for Ae. albopictus and Ae. triseriatus.     

Structures of sialylated oligosaccharides of human erythropoietin expressed in recombinant BHK-21 cells

The native structures of the Asn-linked oligosaccharides and the O-glycans at Ser126 of human erythropoietin expressed from recombinant BHK cells have been elucidated. Enzymatically released N-glycans were studied by methylation analyses, fast-atom-bombardment mass spectrometry as well as one- and two-dimensional 1H-NMR spectrometry at 600 MHz. Many (82.7%) were found to be tetraantennary N-acetyllactosamine-type (22.8% with one, 3.6% with two and 0.4% with three N-acetyllactosamine repeats) being tetrasialylated (41%), trisialylated (29.6%) and disialylated (12.2%). A few (9.7%; 4.1% 2,4-branched, 5.6%, 2,6-branched) of the chains were triantennary (5.4% trisialyl, 4.3% disialyl) and 4.6% were of the disialyl diantennary type. Almost all of the innermost GlcNAc residues were alpha 1-6 fucosylated and NeuAc was exclusively alpha 2-3 linked to Gal beta 1-4GlcNAc-R; 60% of the protein was found to be O-glycosylated at Ser126; structures were monosialylated (70%) or disialylated (30%) forms of the Gal beta 1-3GalNAc core type. Glycosylation patterns at individual Asn-Xaa-Thr/Ser sites were determined by analytical high-pH anion-exchange chromatography with pulsed amperometric detection. Only tetraantennary chains with 0-3 N-acetyllactosamine repeats were detected at Asn38 and Asn83, while almost all of the di- and triantennary oligosaccharides were attached to Asn24. Batch analysis of different preparations of recombinant erythropoietin revealed the high reproducibility of the production procedure. Structures containing terminal GalNAc-GlcNAc were detected in small amounts in a few batches.     

N- and O-glycosylation muteins of recombinant human erythropoietin secreted from BHK-21 cells

Single-site glycomuteins of recombinant human erythropoietin (rhuEpo) were constructed and transiently and stably expressed in BHK-21 cells. The transient expression levels varied among muteins, being highest for mutein rhuEpoGln24 followed by wild-type rhuEpo (rhuEpowt). All other glycomuteins, including rhuEpoGln38, rhuEpoGln83, rhuEpoThr126, and rhuEpoGly126, were secreted at lower levels than rhuEpowt. Muteins expressed in stable cell lines showed similar differences in expression levels. Also each mutein could be affinity-purified from culture supernatants, and was biologically active in vivo. Based on secretion rates from BHK-21 cells, the most potent erythropoietin was rhuEpoGln24. This mutein is also considered to have biologic activities that are superior to rhuEpowt.     

Larval rearing water and preexisting eggs influence oviposition by Aedes aegypti and Ae. albopictus (Diptera: Culicidae)

Two-choice laboratory bioassays were used to evaluate oviposition responses of gravid Aedes aegypti (L.) and Ae. albopictus (Skuse) to larval rearing water and eggs from prior egg deposition. Oviposition responses by female Ae. aegypti were similar between larval water from Ae. aegypti and water controls but significantly greater to larval water from Ae. albopictus (12.2%) compared with water controls. Oviposition by Ae. albopictus females increased significantly in response to larval water from either species compared with water controls with increases of 15.1% to Ae. aegypti larval water and 17.6% to Ae. albopictus larval water. Oviposition responses of gravid Ae. aegypti and Ae. albopictus were evaluated in the presence of preexisting conspecific and heterospecific eggs on strips of oviposition paper. Significantly more eggs were laid by gravid Ae. aegypti females on oviposition paper containing either Ae. aegypti or Ae. albopictus eggs than on oviposition paper without eggs. In contrast, oviposition responses of gravid Ae. albopictus females were unaffected by the presence of eggs of either species.     

Duration of larval and pupal development stages of Aedes albopictus in natural and artificial containers

Mitochondrial function in muscle in vivo can be quantitatively evaluated using 31-phosphorus nuclear magnetic resonance. In resting muscle, the concentrations of ions (e.g. H+, Na+) and two of the major bioenergetic components (inorganic phosphate and creatine) are determined by regulated transcellular transport processes. During recovery after exercise the kinetics and control of mitochondrial ATP synthesis can be established. During exercise the relative contributions to ATP synthesis of phosphocreatine (using creatine kinase), anaerobic glycogenolysis and oxidative phosphorylation are dissected and have been shown to change with time. The consequences of mitochondrial lesions and dysfunctions on these processes have been summarised.